【作者】 武晓玲；

【导师】 迟路湘；

【作者基本信息】 第三军医大学 ， 神经病学， 2006， 硕士

【摘要】 背景和目的颈动脉粥样硬化性狭窄(carotid atherosclerotic stenosis,CASS)是缺血性脑血管病(Ischemic Cerebral Vascular Disease,ICVD)的重要危险因素。造成脑卒中的机制之一是斑块不稳定、破裂,破裂的斑块栓塞远端血管,或斑块表面粗糙,形成血栓。血液的流动会对血管壁不断产生各种力的作用,包括:切应力(剪切力)(Shear stressSs)、周应力(circumferential stress CS)和压应力(pressure stress PS)等。其中剪切应力认为是与动脉粥样硬化发生及斑块破裂最为相关的力学因素。研究表明,血管狭窄程度大于40%可引起血流动力学变化,这种血流动力学改变可影响斑块稳定性。文献认为:狭窄前端过高的剪切应力使血小板源性生长因子(platelet-derived growth factor,PDGF)分泌减少,抑制了血管平滑肌细胞的蛋白合成,细胞凋亡增加,斑块的纤维帽变薄而易于破裂。因此,狭窄动脉局部流体力学因素是动脉粥样硬化具有高度病灶性的原因。有作者认为,脑动脉硬化病变初期,中膜平滑肌细胞分裂增殖,并迁移到内膜,分泌结缔组织基质,包括胶原纤维、弹性纤维和蛋白多糖,引起内膜增厚,参与内膜粥样硬化斑的形成。病变进一步发展,管腔狭窄导致动脉内血流方式发生变化,使正常血管变为弯管,而这种弯管将迫使血流转向,导致血流对管壁产生较大的压力,从而使管壁周应力增大,进一步造成血管壁特别是中膜的损伤,中膜则有不同程度的萎缩。血管壁的力学性质主要取决于中层,后者又取决于其中胶原纤维、弹性纤维和平滑肌的性质、含量及空间构型。动脉管腔局部狭窄不仅使动脉几何形状及力学性质发生显著改变,更严重的是引起动脉中的血液流动和流场异化,诱导动脉组织增生,促使动脉进一步狭窄和更加粥样硬化;并随着狭窄的发展流体力学因素起着越来越大的作用,从而造成严重的循环系统疾病。国外多中心前瞻性研究结果表明,对有严重的颈动脉粥样硬化及狭窄的患者,早期进行颈动脉内膜剥脱术(carotid endarterectomy CEA),可使重度颈动脉狭窄患者的年卒中发病率降低到4.5%。但存在一定的手术死亡及致残率,许多患者难以接受。而且,CEA手术创伤较大,不适合于高危患者及高位狭窄、多发狭窄、颅内动脉狭窄以及椎动脉狭窄等。近年来随着神经影像和神经介入治疗的发展,相继开展了经皮腔内血管成形术(percutaneous transluminal angioplasty PTA)和支架植入术(Stenting)治疗颈动脉狭窄,并针对此种方法的脑栓塞、再狭窄等并发症研制出相应的脑保护装置及防止支架植入后内膜增生的方法,使脑血管介入治疗成为现实。支架置入血管成形术治疗颈动脉狭窄已被公认为有肯定疗效的治疗方法。但其手术适应征仍未达成共识。有学者提出,狭窄程度为30%-69%的有症状或无症状病人是否也可从支架治疗中获益?我们认为:不同狭窄程度血流动力学改变对内、中膜病理形态、管壁弹性特征造成的影响不同,从而影响支架置入效果。因此,本实验目的在于建立最经济、有效的粥样硬化动物模型,研究不同狭窄程度粥样硬化颈动脉流场改变对斑块形态及管壁病理结构、血管力学特性影响,有助于寻找最佳治疗时机,为临床治疗提供理论依据。研究方法:1、采用空气内膜干燥术加高脂饮食建立兔颈动脉内膜损伤及粥样斑块形成模型。2、模型制备成功后,将动物随机分为正常对照组,高脂对照组,病理组及生物力学测试组。3、病理组于术后2、4、8、12周利用彩色血流多普勒超声探测平均血流速度,舒张末期血管内径。同时采集静脉血进行血液流变学检查,测定血液黏度,结合超声所测数据计算血流剪切力。利用数字减影血管造影与计算机图形处理系统进行狭窄的定性与定量研究。留取颈动脉标本做HE染色、Mosson染色和免疫组化染色等,通过计算机图像分析系统计算内-中膜比值,管壁平滑肌及胶原纤维平均积分光密度。高脂组在假手术后2周与正常对照组完成上述测试。4、生物力学测试组于术后2、4、8、12周测定血管弹性模量(Elastic modulus EM)、抗拉强度(Strength failure SF)、最大载荷(peak load PD)等,画出应力-应变曲线(Stress-strain curve SSC)。对比不同狭窄程度流场变化对内中膜形态及血管壁力学特性的影响。研究结果:实验1.各组平均狭窄率分别是:A组为:36.58%±8.53%;B组为:60.28%±5.08%;C组为:76.07%±5.35%;D组为:65.58%±8.36%;75%实验动物颈动脉有小侧支,结扎后干燥效果更好。病理组48根实施手术血管,镜下斑块发生率对比:左侧87.5%,右侧54%,两者比较有差异(P＜0.05),双侧血管重度狭窄比例对比:左侧重度狭窄比例为45%,而右侧重度狭窄比例仅为8.3%,两者比较有差异(P＜0.05)。实验2.手术组颈动脉出现较典型的粥样硬化改变。2周组以纤维斑块为主,中膜明显变厚,平滑肌平均积分光密度值增加,但随狭窄程度增高和剪切力增加,狭窄血管流场改变,4周及8周组出现纤维帽变薄、表面溃破,脂质核心明显增大、斑块内部出血等不稳定斑块特点。血管中膜病理改变以萎缩变薄及平滑肌含量减少为突出特点。统计显示:2周组与4、8、12周组相比均有显著差异(P＜0.05)。免疫组化染色显示,不稳定斑块CD68表达增多,而且狭窄上游区更为显著。实验3.随粥样硬化程度增加,血管内膜增厚,中膜变薄,大量增生的胶原组织代替平滑肌纤维,血管弹性模量增高,硬度增大,顺应性减低。各组间数据比较,12周组与高脂组、手术后2、4、8周组相比均有显著差异。但2周组与高脂组对比差异不明显。结论:1.经过对实验方法的简单改进,我们利用高脂饮食加空气干燥术成功复制出兔颈动脉粥样硬化模型,部分斑块显示出不稳定斑块特点,有利于对粥样硬化病理机制的更深入研究。2.我们发现:(1)、75%兔颈总动脉存在细小血管分支。术中仅对左侧血管分支暂时结扎,双侧手术后结果对比左侧干燥效果更好,成模率更高。(2)本实验成功的在双侧兔颈动脉实施空气干燥手术,未发现增加手术死亡率。3.随着血管狭窄程度增高,血流剪切力增加、湍流形成、狭窄血管流场改变,斑块稳定性下降,血管发生重构改变。2周组以平滑肌增殖为主,而4、8、12周组表现为胶原增生,中膜萎缩变薄,斑块显示不稳定特征。4.在动脉粥样硬化早期,血管呈正性重塑,弹性模量略有降低以适应新的力学环境。但随着粥样硬化程度加重,动脉壁胶原含量增加,内膜增厚而中膜萎缩变薄,管壁弹性减低,硬度增加,狭窄后段扩张且易形成动脉夹层,为介入治疗增加难度。更多还原

【Abstract】 Backgroud and objective:Carotid atherosclerotic stenosis（CASS） is an important risk factor of ischemic Cerebra Vascular Disease（ICVD）.One of the mechanism of stroke is vulnerable and rupturable plaque which embolize the distal vessel,or the rough plaque surface form the embolism.The flow of blood can bring the various kinds action of force,that include shear stress, circumferential stress,pressure stress so on.The shear stress is considered that it is a mechanics parameter that the best correlative with the formation of the atherosclerosis and rupture of the plaque.According to the research,that the degree of vessel stenosis exceed 40%can bring dynamics changs.This dynamics changs can influence the stability of the plaque.Literature shows,that the exorbitant shear stress at the upstream of stenosis decrease the secretion of the PDGF and restrained the protein compose of the vessel smooth muscle cell（SMC）,meanwhile the cell apoptosis increase,the fibrous cap become thin and tend rupture.So,the hydrodynamics factors in the local of the stenotic artery is reason of the atherosclerosis which show the high pathogenicity.Some author consider that,in the initial stage of brain atherosclerosis,the media SMC division and proliferation,transfer to the intima,secrete the connective tissue matrix, include the collagen fibers,elastic fiber and proteoglycan,to cause the intima thickening, and participate the form of As plaque.Following the development of disease,the vessel lumina stenosis result in the blood flow fashion change and the normal vessel become the bend lumina,which will change direction of blood flow and bring the large pressure strain in the vessel wall by blood flow,thus the circumferential stress increase,vessel wall especially at media injury,media occours atrophy in different degrees.The mechanics character of the vessel wall lie on the media,but the media lie on the character,content and spatial configuration of the collagen fibers,elastic fiber and the SM fiber.artery lumina of the local stenosis not only significantly change the geometry and mechanics character of artery,but also bring blood flow and flow field dissimilation, derivation artery hyperblastosis,and accelerate artery stenosis and As.Then following the development of stenosis,the hydrodynamics factors bring the more and more effect.and result in the severe recirculating system disease.Overseas multicentre prospective research manifested,that the patient with the severe carotid atherosclerotic and stenosis was performed the CEA in the earlier period,can decrease the morbidity of the stroke in a year to the 4.5%.But it bring a number of disbility rate and the death rate,thus most patient unwillingness to accept CEA.Furthermore CEA may bring the major trauma,and it is unsuitable with the high risk and perch stenosis,multi-stenosis,Intracranial and vertebral artery stenosis.Recently following the development of the neuroimaging and the neuro interventional therapy,PTA and stenting has was one after another carried out to therapy CASS,and the brain protective device was manufactured to aim at preventing the cerebral embolism and restenosis,thus the cerebral vascular interventional therapy become reality.PTAS to treat the CASS was generally accepted as a therapy of positive curative effect. But its indcation can not be identification.Some scholar propose,if can the patient of symptomatic or asymptomatic of degree of stenosis 30-69%benefit from the interventional therapy? We think,the change of hemodynamics in the different degree stenosis carotid artery bring different effect on intimia-media pathological and the mechanics parameter changes,thus effect the result of interventional therapy.Accordingly, our experiment has studied that the changes of the flow field in the atherosclerotic carotid artery of rabbit and its effect on intimia-media pathology and the mechanics parameter,conduce to search the optimization treatment opportunity.Methods:1.To establish carotid artery intimia lesion and atherosclerosis model by high cholesterol diet Plus airdrying.2.Japanese White rabbits were randomly divided into blank control group（n=6）,sham operation control group（n=6）,the pathology group（n=24） and mechanics test group.3.At 2^th、4^th、8^th and 12^th week after operation,the degree of mean blood flow velocity and diastasis vessel inner diameter were measured using Color Doppler Flow Imaging to the pathology group.Meanwhile the vein blood was collected and blood viscosity was measured,the shear stress was calculated by CDFI measuring datum.The degree of artery stenosis was analyzed qualitatively and quantitatively by DSA and computer graphic processing system.Carotid artery samples were reserved and the HE,Masson and immunohistochemically was carried out.Intimia-media ratio,mean integra optical densities（IOD） of media smooth muscle cell（SMC） and collagen were calculated. The control groups were measured after 2^th.4.The mechanical properties were measured on a tensile-testing machine to the mechanical test group.The Elastic modulus,the strength failure and the peak load was measured.stress-strain curve were drawn.The change of different degree of stenosis flow field and intimia-media pathological,the vessel wall mechanics parameter changes were compared.Results:1.The average rates of stenosis were calculated and described as following,2-week group:36.58%±8.53%;4-week group 60.28%±5.08%;8-week group 76.07%±5.35%; 12-week65.58%±8.36%;75%carotid arteries of rabbit have the little ramus.The effect was more satisfying after the ramus been ligated temporarily.48 surgery vessel,incidence rate of plaque by microscope checking up:left 87.5%,right 54%,result showed significant difference.,The rates of severe stenosis of bilateral vessel:left 45%,right 8.3%showed significant difference..2.The typical atherosclerotic plaque showed in surgery group.The fibrous plaque was observed,mean while thickness of media and mean IOD of SMC increased at 2^th week after operation.However,following the increase of the stenosis rate and shear stress,the flow field was changed on the stenotic artery.The pathological morphology showed the character of vulnerable plaque such a large lipid core,thin fibrous cap,and rupture in the shouder of plaque at 4^th 8^th week after operation.The media showed atrophy and became thin,mean IOD of SMC was decreased.Statistic analysis showed significant difference in the 2 week group,compared with other group.Immunohistochemically show,vulnerable plaque CD68 express increase.,and upstream region of the plaque is even more obvious.3.Following degrees of the atherosclerosis,the intima showed proliferation and media showed atrophy and became thin,a mass of collagen tissue proliferated and filled in the smooth muscle cell.Artery elastic modulus and stiffness showed increase and atery compliance showde decrease.Statistic analysis showed significant difference at the 12^th week group,compared with sham operation control group and 2^th 4^th 8^th week group after operation,but it showed no such significant deviations at 2^th week group after operation,compared with sham operation control groupConclusions:1.Through the several improvement on the experiment method,we successfully established a carotid atherosclerosis model induced by cholesterol diet plus air-drying in the rabbit.The part of plaque showed the vulnerable character.The pathological changes and its mechanisms were more suitable for the future research in experimental carotid atherosclerosis.2.75%carotid arteries of rabbit have the little ramus.The effect was more satisfying after the ramus been ligated temporarily.Bilateral carotid arteries of rabbit were treated,but only ramus of left carotid arteries was ligated temporarily.The significant result was observed in the left carotid artery,without increase of mortality.3.Following the degree of stenosis,the flow shear stress increased,turbulent flow emerged and flow field changed at the stenotic artery.The plaque become unstable.The vessel occured remodeling.4.At the early atherosclerotic carotid artery,vessel showed positive remodeling.elastic modulus decreased to adapt to the new mechanical condition.But following degrees of the atherosclerosis,.the content of the artery wall collagen increased,the vessel wall elasticity decreased and stiffness increased.The vessel occured negative remodeling,which suggested that the intervetional treatment be in difficulty.更多还原